Antibody-Drug-Conjugates (ADC's) which are useful for the treatment of cancer and other diseases are commonly composed of three distinct elements: a cell-binding agent; a linker; and a cytotoxic agent. The conventional method of conjugating a cell-binding agent, such as an antibody, to a cytotoxic agent, employs two distinct reaction steps. In the first reaction step, the antibody is reacted with a bifunctional crosslinking reagent to produce a linker-modified antibody. The modified antibody product is then optionally purified from the excess linker or hydrolyzed linker reagent. In the second reaction step, the linker-modified antibody is reacted with the cytotoxic agent containing a reactive group, such as thiol, to generate the antibody-cytotoxic agent conjugate, which is again purified in an additional purification step. This conventional method may require two purification steps, which lowers the overall yield and makes the process cumbersome and uneconomical for large scale manufacturing of ADCs.
Alternatively, the cytotoxic agent can first react with the bifunctional crosslinking reagent to form a cytotoxic agent-linker compound, which is then reacted with a cell-binding agent to form the cell-binding agent-cytotoxic agent conjugate (conjugation reaction). The cytotoxic agent, the bifunctional crosslinking reagent and the resulting cytotoxic agent-linker compound are generally hydrophobic compounds and have low solubility in water. As such, a certain amount of organic solvent is required to be present in the conjugation reaction to solubilize the cytotoxic agent-linker compound. On the other hand, the presence of large amount of organic solvent may have a detrimental effect on the antibody stability. Accordingly, the process of the art may be limited to small scale preparation of the cell-binding agent-cytotoxic agent conjugates when low concentrations of cell-binding agents, cytotoxic agents and/or bifunctional crosslinking reagents are used.
In view of the foregoing, there is a need to develop improved processes for preparing cell-binding agent-cytotoxic agent conjugates with high antibody stability and are suitable for large scale productions.